Mapping the Distribution of Conformational Information Throughout a Protein Sequence

Departamento de Ciencia y Tecnología, Universidad Nacional de Quilmes, Roque Sáenz Peña 180, (1876) Bernal, Buenos Aires, Argentina.
Journal of Molecular Biology (Impact Factor: 4.33). 05/2006; 358(1):280-8. DOI: 10.1016/j.jmb.2006.01.095
Source: PubMed


The three-dimensional structure of protein is encoded in the sequence, but many amino acid residues carry no essential conformational information, and the identity of those that are structure-determining is elusive. By circular permutation and terminal deletion, we produced and purified 25 Bacillus licheniformis beta-lactamase (ESBL) variants that lack 5-21 contiguous residues each, and collectively have 82% of the sequence and 92% of the non-local atom-atom contacts eliminated. Circular dichroism and size-exclusion chromatography showed that most of the variants form conformationally heterogeneous mixtures, but by measuring catalytic constants, we found that all populate, to a greater or lesser extent, conformations with the essential features of the native fold. This suggests that no segment of the ESBL sequence is essential to the structure as a whole, which is congruent with the notion that local information and modular organization can impart most of the tertiary fold specificity and cooperativity.

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    • "However, bioassays are, in general, expensive, complex, and not always available in a research or biotechnology laboratory (see below). An approach to study the functional activity of the protein would be to characterize its enzymatic activity [11], if any, or its interaction with a specific ligand, which is usually the first step in protein-induced cellular response . On the other hand, there is a battery of biophysical methods, some of them simple and inexpensive that are highly useful for testing conformation of every protein product (Table 1). "
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    ABSTRACT: Production of recombinant proteins is a process intensively used in the research laboratory. In addition, the main biotechnology market products are recombinant proteins and monoclonal antibodies. The biological (and clinical) properties of the protein product strongly depend on the conformation of the polypeptide. Therefore, assessment of the correct conformation of the produced protein is crucial. There is no single method to assess every aspect of protein structure or function. Depending on the protein, the methods of choice vary. There are general methods to evaluate not only mass and primary sequence of the protein, but also higher-order structure. This review outlines the principal techniques for determining the conformation of a protein from structural (biophysical methods) to functional (in vitro binding assays) analyses.
    Biotechnology Journal 06/2011; 6(6):731-41. DOI:10.1002/biot.201100107 · 3.49 Impact Factor
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    • "The outcomes of these previous studies have indicated that CPs usually retain native structures and biological functions (3–5,9,10), although the stabilities and folding mechanisms might be altered (7,11,12). Since CP may sometimes increase the stability (13), activity or functional diversity (14–16) of proteins, it has been applied to trigger crystallization (13), improve enzyme activities (14), determine critical elements (17,18) and create novel fusion proteins (19–22). "
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    ABSTRACT: Circular permutation (CP) in a protein can be considered as if its sequence were circularized followed by a creation of termini at a new location. Since the first observation of CP in 1979, a substantial number of studies have concluded that circular permutants (CPs) usually retain native structures and functions, sometimes with increased stability or functional diversity. Although this interesting property has made CP useful in many protein engineering and folding researches, large-scale collections of CP-related information were not available until this study. Here we describe CPDB, the first CP DataBase. The organizational principle of CPDB is a hierarchical categorization in which pairs of circular permutants are grouped into CP clusters, which are further grouped into folds and in turn classes. Additions to CPDB include a useful set of tools and resources for the identification, characterization, comparison and visualization of CP. Besides, several viable CP site prediction methods are implemented and assessed in CPDB. This database can be useful in protein folding and evolution studies, the discovery of novel protein structural and functional relationships, and facilitating the production of new CPs with unique biotechnical or industrial interests. The CPDB database can be accessed at
    Nucleic Acids Research 11/2008; 37(Database issue):D328-32. DOI:10.1093/nar/gkn679 · 9.11 Impact Factor
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